Elusive data underlying debate at the prokaryote-eukaryote divide

Background

The origin of eukaryotic cells was an important transition in evolution. The factors underlying the origin and evolutionary success of the eukaryote lineage are still discussed. One camp argues that mitochondria were essential for eukaryote origin because of the unique configuration of internalized bioenergetic membranes that they conferred to the common ancestor of all known eukaryotic lineages. A recent paper by Lynch and Marinov concluded that mitochondria were energetically irrelevant to eukaryote origin, a conclusion based on analyses of previously published numbers of various molecules and ribosomes per cell and cell volumes as a presumed proxy for the role of mitochondria in evolution. Their numbers were purportedly extracted from the literature.

Results

We have examined the numbers upon which the recent study was based. We report that for a sample of 80 numbers that were purportedly extracted from the literature and that underlie key inferences of the recent study, more than 50% of the values do not exist in the cited papers to which the numbers are attributed. The published result cannot be independently reproduced. Other numbers that the recent study reports differ inexplicably from those in the literature to which they are ascribed. We list the discrepancies between the recently published numbers and the purported literature sources of those numbers in a head to head manner so that the discrepancies are readily evident, although the source of error underlying the discrepancies remains obscure.

Conclusion

The data purportedly supporting the view that mitochondria had no impact upon eukaryotic evolution data exhibits notable irregularities. The paper in question evokes the impression that the published numbers are of up to seven significant digit accuracy, when in fact more than half the numbers are nowhere to be found in the literature to which they are attributed. Though the reasons for the discrepancies are unknown, it is important to air these issues, lest the prominent paper in question become a point source of a snowballing error through the literature or become interpreted as a form of evidence that mitochondria were irrelevant to eukaryote evolution.

Reviewers

This article was reviewed by Eric Bapteste, Jianzhi Zhang and Martin Lercher.

     

Energetics and dynamics of SNAREpin folding across lipid bilayers

Membrane fusion occurs when SNAREpins fold up between lipid bilayers. How much energy is generated during SNAREpin folding and how this energy is coupled to the fusion of apposing membranes is unknown. We have used a surface forces apparatus to determine the energetics and dynamics of SNAREpin formation and characterize the different intermediate structures sampled by cognate SNAREs in the course of their assembly. The interaction energy-versus-distance profiles of assembling SNAREpins reveal that SNARE motifs begin to interact when the membranes are 8 nm apart. Even after very close approach of the bilayers (approximately 2-4 nm), the SNAREpins remain partly unstructured in their membrane-proximal region. The energy stabilizing a single SNAREpin in this configuration (35 k(B)T) corresponds closely with the energy needed to fuse outer but not inner leaflets (hemifusion) of pure lipid bilayers (40-50 k(B)T).     

Native oak chloroplasts reveal an ancient divide across Europe

Glacial refugia and postglacial migration are major factors responsible for the present patterns of genetic variation we see in natural populations. Traditionally postglacial history has been inferred from fossil data, but new molecular techniques permit historical information to be gleaned from present populations. The chloroplast tRNA^Leu1 intron contains regions which have been highly conserved over a billion years of chloroplast evolution. Surprisingly, in one of these regions which has remained invariant for all photosynthetic organisms so far studied, we have found intraspecific site polymorphism. This polymorphism occurs in two European oaks, Quercus robur and Q. petraea , indicating hybridisation and introgression between them. Two distinct chloroplast types occur and are distributed geographically as eastern and western forms suggesting that these oaks are each derived from at least two separate glacial refugia.     

The prokaryote-eukaryote dichotomy: meanings and mythology.

Drawing on documents both published and archival, this paper explains how the prokaryote-eukaryote dichotomy of the 1960s was constructed, the purposes it served, and what it implied in terms of classification and phylogeny. In doing so, I first show how the concept was attributed to Edouard Chatton and the context in which he introduced the terms. Following, I examine the context in which the terms were reintroduced into biology in 1962 by Roger Stanier and C. B. van Niel. I study the discourse over the subsequent decade to understand how the organizational dichotomy took on the form of a natural classification as the kingdom Monera or superkingdom Procaryotae. Stanier and van Niel admitted that, in regard to constructing a natural classification of bacteria, structural characteristics were no more useful than physiological properties. They repeatedly denied that bacterial phylogenetics was possible. I thus examine the great historical irony that the "prokaryote," in both its organizational and phylogenetic senses, was defined (negatively) on the basis of structure. Finally, we see how phylogenetic research based on 16S rRNA led by Carl Woese and his collaborators confronted the prokaryote concept while moving microbiology to the center of evolutionary biology.     

Discord in the family Sparidae (Teleostei): divergent phylogeographical patterns across the Atlantic-Mediterranean divide

The Strait of Gibraltar has been proposed to be the divide between two marine biogeographical regions, the Mediterranean Sea and the Northeast Atlantic. Intraspecific studies have shown, for several of the examined species, a reduction of gene flow between the two basins. The present study examines genetic variation at nuclear and mitochondrial loci in five marine teleost species belonging to the family Sparidae. Four samples for each species were analysed spanning the Northeast Atlantic and the Mediterranean. For all individuals 17 allozyme loci were scored and a combined single strand conformation polymorphism-sequencing approach was used to survey approximately 190 bp of the mitochondrial DNA (mtDNA) D-loop region. All five species share similar biological features. For three species, namely Lithognathus mormyrus, Spondyliosoma cantharus, and Dentex dentex, large mtDNA divergence was observed between Atlantic and Mediterranean samples. Little or no mtDNA differentiation was found in the other two species, Pagrus pagrus and Pagellus bogaraveo. Allozyme data revealed strong differentiation when comparing Atlantic and Mediterranean samples of L. mormyrus and D. dentex, moderate for P. pagrus, and no differentiation for P. bogaraveo and S. cantharus. These results provide evidence for a sharp phylogeographical break (sensu Avise) between the Atlantic and the Mediterranean for two (or possibly three) sparid species of the five investigated. At the same time, the obtained results for the other two species raise the question on which ecological/historical factors might have caused the observed discrepancy in the geographical distribution of genetic variation among otherwise biologically similar species.     

Differences in timing of migration and response to sexual signalling drive asymmetric hybridization across a migratory divide

Ecological traits and sexual signals may both contribute to the process of ecological speciation. Here we investigate the roles of an ecological trait, seasonal migratory behaviour and a sexual trait, song, in restricting or directing gene flow across a migratory divide in the Swainson's thrush (Catharus ustulatus). We show that short-distance migratory ecotypes wintering in Central America arrive earlier at the breeding grounds than long-distance migratory ecotypes wintering primarily in South America, providing the potential for some premating isolation. Playback experiments suggest that early- and late-arriving forms recognize each other as competitors, but that the early-arriving form responds more aggressively to a broader spectrum of stimuli. Genetic analysis suggests that hybridization occurs more often between males of the early-arriving ecotype and females of the late-arriving ecotype. Together our results suggest that differences in arrival times may reduce the temporal coincidence of mate choice, but asymmetry in response to heterotypic song may hinder complete divergence. These data provide further insight into the roles of ecological traits and sexual signals during the incipient stages of speciation.     

Energetics of protein transport across biological membranes. a study of the thylakoid DeltapH-dependent/cpTat pathway

Among the pathways for protein translocation across biological membranes, the DeltapH-dependent/Tat system is unusual in its sole reliance upon the transmembrane pH gradient to drive protein transport. The free energy cost of protein translocation via the chloro-plast DeltapH-dependent/Tat pathway was measured by conducting in vitro transport assays with isolated thylakoids while concurrently monitoring energetic parameters. These experiments revealed a substrate-specific energetic barrier to cpTat-mediated transport as well as direct utilization of protons from the gradient, consistent with a H+/protein antiporter mechanism. The magnitude of proton flux was assayed by four independent approaches and averaged 7.9 x 10(4) protons released from the gradient per transported protein. This corresponds to a DeltaG transport of 6.9 x 10(5) kJ.mol protein translocated(-1), representing the utilization of an energetic equivalent of 10(4) molecules of ATP. At this cost, we estimate that the DeltapH-dependent/cpTat pathway utilizes approximately 3% of the total energy output of the chloroplast.     

Survival of the currently fittest: genetics of rainbow trout survival across time and space

As a fitness trait, survival is assumed to exhibit low heritability due to strong selection eroding genetic variation and/or spatio-temporal variation in mortality agents reducing genetic and increasing residual variation. The latter phenomenon in particular may contribute to low heritability in multigeneration data, even if certain cohorts exhibit significant genetic variation. Analysis of survival data from 10 year classes of rainbow trout reared at three test stations showed that treating survival as a single trait across all generations resulted in low heritability (h(2) = 0.08-0.17). However, when heritabilities were estimated from homogeneous generation and test station-specific cohorts, a wide range of heritability values was revealed (h(2) = 0.04-0.71). Of 64 genetic correlations between different cohorts, 20 were positive, but 16 were significantly negative, confirming that genetic architecture of survival is not stable across generations and environments. These results reveal the existence of hidden genetic variation for survival and demonstrate that treating survival as one trait over several generations may not reveal its true genetic architecture. Negative genetic correlations between cohorts indicate that overall survival has limited potential to predict general resistance, and care should be taken when using it as selection criterion.     

Widespread and structured distributions of blood parasite haplotypes across a migratory divide of the Swainson's thrush (Catharus ustulatus)

We examined the phylogenetic distribution of cytochrome b haplotypes of the avian blood parasite genera Haemoproteus and Plasmodium across the migratory divide of the Swainson's thrush (Catharus ustulatus) in British Columbia, Canada. From 87 host individuals, we identified 8 parasite haplotypes; 4 of Plasmodium and 4 of Haemoproteus. Six haplotypes were novel; 1 Haemoproteus haplotype was identical to H. majoris found in the blue tit (Parus caeruleus) in Sweden, and another halotype was identical to a Plasmodium haplotype found in the white-crowned sparrow (Zonotrichia leucophrys) in Oregon. The 2 most abundant parasite haplotypes were widely distributed across the contact zone, whereas 2 other parasite haplotypes seem to have structured distributions. Compared with 74 Plasmodium and Haemoproteus haplotypes published in GenBank, haplotypes recovered from Swainson's thrushes do not form monophyletic groups, and they are closely related to haplotypes from a variety of other hosts and localities. In addition, we recovered 2 Swainson's thrush Plasmodium haplotypes from the nonmigratory orange-billed nightingale thrush (Catharus aurantiirostris) in Costa Rica. This study is the first to elucidate avian blood parasite transmission, distribution, and phylogenetic relationships in an avian contact zone in North America.     

Muscle Energetics and the Fenn Effect

This study suggests and statistically tests a consistent analytical method for isolating the critical variables governing the Fenn effect in frog sartorius muscle. It demonstrates that when the Hill factor (P₀ - P) is used to normalize the heat of shortening, or the work, or their sum, the enthalpy of shortening, these reduced energy terms are highly linear functions of the time of shortening. Linear correlation of a given form of reduced energy for a given muscle, against the time of shortening yields correlation coefficients of 0.998 or greater. From the regression equations, equations similar in form to the 1938 Hill force-velocity, and 1964 Hill heat of shortening equations, are deduced. The analysis suggests that the efficiency with which the extra energy of shortening is converted into work is nearly constant over most of the range of fractional loads, and it confirms Fenn's observation that the total extra energy of shortening is about 1.3 times the work. Thus it is also consistent with the results of most of the biochemical studies which correlate the extra breakdown of high energy phosphates entirely with the work done, and no component exclusively with the distance shortened. While the analysis successfully identifies the two factors regulating the release of the energy of shortening, time and the fractional load, there was insufficient data in the literature to develop the form of the relation between the size and the geometry of the muscle and the rate of energy release. Therefore until more data are available it seems unjustified to assume that either b/l₀ and a/P₀ (or the equivalent linear regression constants) are independent of these factors.     

Soil nitrogen availability varies with plant genetics across diverse river drainages

Understanding covariance of plant genetics and soil processes may improve our understanding the role of plant genetics in structuring soils and ecosystem function across landscapes. We measured soil nitrogen (N) and phosphorus (P) availability using ion exchange resin bags within three river drainages across Utah and Arizona, USA. The three drainages spanned more than 1,000 km in distance, 8° of latitude, and varying climatic regimes, but were similarly dominated by stands of Populus fremontii (S. Watts), P. angustifolia (James), or natural hybrids between the two species. Soil N availability was consistently greater in P. fremontii stands compared to P. angustifolia stands, and hybrid stands were intermediate. However, we found that the influence of overstory type on soil P availability depended on the river drainage. Our study suggests that, even with a near doubling of mean soil N availability across these drainages, the relative genetic-based effects of the dominant plant on N availability remained consistent. These results expand upon previous work by: 1) providing evidence for linkages between plant genetic factors and ecosystem function across geographic scales; and 2) indicating that plant genetic-based effects on nutrient dynamics in a given ecosystem may differ among nutrients (e.g., N vs. P).     

Integrative approach to pain genetics identifies pain sensitivity loci across diseases

Identifying human genes relevant for the processing of pain requires difficult-to-conduct and expensive large-scale clinical trials. Here, we examine a novel integrative paradigm for data-driven discovery of pain gene candidates, taking advantage of the vast amount of existing disease-related clinical literature and gene expression microarray data stored in large international repositories. First, thousands of diseases were ranked according to a disease-specific pain index (DSPI), derived from Medical Subject Heading (MESH) annotations in MEDLINE. Second, gene expression profiles of 121 of these human diseases were obtained from public sources. Third, genes with expression variation significantly correlated with DSPI across diseases were selected as candidate pain genes. Finally, selected candidate pain genes were genotyped in an independent human cohort and prospectively evaluated for significant association between variants and measures of pain sensitivity. The strongest signal was with rs4512126 (5q32, ABLIM3, P = 1.3×10⁻¹⁰) for the sensitivity to cold pressor pain in males, but not in females. Significant associations were also observed with rs12548828, rs7826700 and rs1075791 on 8q22.2 within NCALD (P = 1.7×10⁻⁴, 1.8×10⁻⁴, and 2.2×10⁻⁴ respectively). Our results demonstrate the utility of a novel paradigm that integrates publicly available disease-specific gene expression data with clinical data curated from MEDLINE to facilitate the discovery of pain-relevant genes. This data-derived list of pain gene candidates enables additional focused and efficient biological studies validating additional candidates.     

Evidence for a freezing tolerance-growth rate trade-off in the live oaks (Quercus series Virentes) across the tropical-temperate divide

? It has long been hypothesized that species are limited to the north by minimum temperature and to the south by competition, resulting in a trade-off between freezing tolerance and growth rate. We investigated the extent to which the climatic origins of populations from four live oak species (Quercus series Virentes) were associated with freezing tolerance and growth rate, and whether species fitted a model of locally adapted populations, each with narrow climatic tolerances, or of broadly adapted populations with wide climatic tolerances. ? Acorns from populations of four species across a tropical-temperate gradient were grown under common tropical and temperate conditions. Growth rate, seed mass, and leaf and stem freezing traits were compared with source minimum temperatures. ? Maximum growth rates under tropical conditions were negatively correlated with freezing tolerance under temperate conditions. The minimum source temperature predicted the freezing tolerance of populations under temperate conditions. The tropical species Q. oleoides was differentiated from the three temperate species, and variation among species was greater than among populations. ? The trade-off between freezing tolerance and growth rate supports the range limit hypothesis. Limited variation within species indicates that the distributions of species may be driven more strongly by broad climatic factors than by highly local conditions.     

Geographic distribution and genetics of killer phenotypes for the yeast Pichia kluyveri across the United States.

Representative strains (n = 61) of the yeast Pichia kluyveri from across the United States were studied for their ability to kill 71 other strains (representing 25 species) of yeast. This survey showed killing activity in 69% of the P. kluyveri strains tested. More extensive analysis of killer activity of 197 P. kluyveri strains against strains of five tester species showed comparable activity (67% of strains tested). This activity was shown to be equally variable within localities, within regions, and across the continent. The genetic basis of the variability was ascertained by tetrad analysis and is most likely due to alleles segregating at three epistatic loci. Evidence for the idea that killer toxins have a role in excluding other yeasts from particular habitats is discussed.     

Geographic distribution and genetics of killer phenotypes for the yeast Pichia kluyveri across the United States.

Representative strains (n = 61) of the yeast Pichia kluyveri from across the United States were studied for their ability to kill 71 other strains (representing 25 species) of yeast. This survey showed killing activity in 69% of the P. kluyveri strains tested. More extensive analysis of killer activity of 197 P. kluyveri strains against strains of five tester species showed comparable activity (67% of strains tested). This activity was shown to be equally variable within localities, within regions, and across the continent. The genetic basis of the variability was ascertained by tetrad analysis and is most likely due to alleles segregating at three epistatic loci. Evidence for the idea that killer toxins have a role in excluding other yeasts from particular habitats is discussed.     

Network-level and population genetics analysis of the insulin/TOR signal transduction pathway across human populations

Genes and proteins rarely act in isolation, but they rather operate as components of complex networks of interacting molecules. Therefore, for understanding their evolution, it may be helpful to take into account the interaction networks in which they participate. It has been shown that selective constraints acting on genes depend on the position that they occupy in the network. Less understood is how the impact of local adaptation at the intraspecific level is affected by the network structure. Here, we analyzed the patterns of molecular evolution of 67 genes involved in the insulin/target of rapamycin (TOR) signal transduction pathway. This well-characterized pathway plays a key role in fundamental processes such as energetic metabolism, growth, reproduction, and aging and is involved in metabolic disorders such as obesity, insulin resistance, and diabetes. For that purpose, we combined genotype data from worldwide human populations with current knowledge of the structure and function of the pathway. We identified the footprint of recent positive selection in nine of the studied genomic regions. Most of the adaptation signals were observed among Middle East and North African, European, and Central South Asian populations. We found that positive selection preferentially targets the most central elements in the pathway, in contrast to previous observations in the whole human interactome. This observation indicates that the impact of positive selection on genes involved in the insulin/TOR pathway is affected by the pathway structure.     

Energetics of the calcium-transporting ATPase

A thermodynamic cycle for catalysis of calcium transport by the sarcoplasmic reticulum ATPase is described, based on equilibrium constants for the microscopic steps of the reaction shown in Equation 1 under a single set of experimental (formula; see text) conditions (pH 7.0, 25 degrees C, 100 mM KCl, 5 mM MgSO4): KCa = 5.9 X 10(-12) M2, K alpha ATP = 15 microM, Kint = 0.47, K alpha ADP = 0.73 mM, K'int = 1.7, K"Ca = 2.2 X 10(-6) M2, and Kp = 37 mM. The value of K"Ca was calculated by difference, from the free energy of hydrolysis of ATP. The spontaneous formation of an acylphosphate from Pi and E is made possible by the expression of 12.5 kcal mol-1 of noncovalent binding energy in E-P. Only 1.9 kcal mol-1 of binding energy is expressed in E X Pi. There is a mutual destabilization of bound phosphate and calcium in E-P X Ca2, with delta GD = 7.6 kcal mol-1, that permits transfer of phosphate to ADP and transfer of calcium to a concentrated calcium pool inside the vesicle. It is suggested that the ordered kinetic mechanism for the dissociation of E-P X Ca2, with phosphate transfer to ADP before calcium dissociation outside and phosphate transfer to water after calcium dissociation inside, preserves the Gibbs energies of these ligands and makes a major contribution to the coupling in the transport process. A lag (approximately 5 ms) before the appearance of E-P after mixing E and Pi at pH 6 is diminished by ATP and by increased [Pi]. This suggests that ATP accelerates the binding of Pi. The weak inhibition by ATP of E-P formation at equilibrium also suggests that ATP and phosphate can bind simultaneously to the enzyme at pH 6. Rate constants are greater than or equal to 115 s-1 for all the steps in the reaction sequence to form E-32P X Ca2 from E-P, Ca2+ and [32P]ATP at pH 7. E-P X Ca2 decomposes with kappa = 17 s-1, which shows that it is a kinetically competent intermediate. The value of kappa decreases to 4 s-1 if the intermediate is formed in the presence of 2 mM Ca2+. This decrease and inhibition of turnover by greater than 0.1 mM Ca2+ may result from slow decomposition of E-P X Ca3.